Summary of work: Cell cycle progression is subject to numerous checkpoints, which serve to arrest growth if certain processes are incomplete, or damage is incurred. While treatment of cells with genotoxic agents often leads to activation of one or more of these checkpoints and results in growth arrest, in some instances such treatment leads to apoptosis. The factors responsible for determining whether a cell will undergo apoptosis or growth arrest have yet to be clarified and this is the focus of this project. Several model systems are being employed to address these questions. In the first, we are investigating the ability of TGFbeta to prevent serum deprivation-induced cell death in human lung carcinoma cells. TGFbeta can prevent the apoptosis of A549 cells that normally occurs following serum withdrawal. Protection is associated with the early induction of c-jun expression, but a later suppression of c-jun-N-terminal kinase (JNK) activity. Expression of dominant negative c-jun protein prevents the protective influence of TGFbeta while expression of a dominant negative form of the kinase SEK1, which is necessary for JNK activation, mimics the effect of TGFbeta. These findings suggest that early expression of c-jun is important for survival as is inhibition of JNK activity at later time points. A second study has focused on the mechanisms contributing to Taxol-induced cell death in MCF-7 cells. We have provided evidence that Taxol induces apoptosis in MCF-7 cells through a caspase-independent mechanism and can be prevented by co-treatment with the serine protease inhibitor TPCK. TPCK treatment also prevents phosphorylation of Raf-1 and Bcl-2 (previously shown to be linked to Taxol-induced cell death). Thus, a TPCK inhibitable protease is required for Taxol-induced apoptosis and functions upstream of Raf-1and Bcl-2phosphorylation in the death pathway. A third project is examining the role for c-jun-N-terminal kinase 2 (JNK2) in tumor cell growth using an antisense strategy to selectively prevent JNK1 and JNK2 expression and activites. We have observed that inhibition of JNK2 expression leads to marked growth inhibition and apoptosis of certain cancer cell lines. Preliminary findings suggest that this effect appears to be restricted to cells lacking p53 function. Current studies are addressing the mechanisms contributing to this effect.